Designing delivery agents for therapeutics is an ongoing challenge. Cell penetrating peptides mimics (CPPMs) represent a class of molecules that can serve as efficient delivery vehicles. Much like their CPP counterparts, CPPMs are inspired by proteins with translocation abilities, such as HIV-1 Tat and Antennapedia homeodomain protein. CPPMs use a synthetic scaffold to capture key features of these proteins critical for cellular uptake, such as cationic charge content, included through the use of arginine-inspired components (guandinium groups), and partial hydrophobicity, through the use of phenylalanine-inspired components (aromatic groups). Here we document efforts to tune CPPM structures for improved delivery of siRNA. Two series of CPPMs with increasing cationic charge contents (10, 20, 40, 60, 120 positive charges) were designed with and without a fixed hydrophobic component to assess the role of carrier composition on delivery efficiencies. FITC-siRNA delivery to Jurkat T cells revealed that CPPMs with 40 charges provided the most efficient delivery, where CPPMs with added hydrophobicity outperformed their less hydrophobic counterparts. When charge content was further increased, delivery efficiency diminished, mirroring a trend from the peptide literature for polyarginine. Promising CPPMs from this series were also shown to outperform common commercially available controls. Efficient delivery of biologically active siRNA was also demonstrated using peripheral blood mononuclear cells (PMBCs), which are traditionally difficult to transfect. Specifically, 50-75% knockdown of the highly regulated gene <i>hNOTCH1 </i>in PMBCs was achieved. By successfully mimicking the function of CPPs we gained insight into the important structural components for efficient delivery and demonstrated the necessary design principles for optimization of these vehicle structures for application-specific needs.

Building new protein mimics for siRNA delivery

249th National Meeting (2015)

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